Triplex-directed and other oligonucleotide-based strategies will be tested for feasibility as gene-specific therapies for autosomal dominant retinitis pigmentosa (ADRP) caused by defects in the rhodopsin gene. In the course of this work the investigator will develop technologies that can be used for testing a variety of therapeutic approaches to ADRP and other autosomal dominant genetic disorders. As any therapy for ADRP must ultimately be tested and optimized in animals, the investigator has chosen mice because of the ease with which their genomes can be modified. The investigator proposes to create a mouse ES cell line selectable segment of DNA that can be efficiently targeted by site-specific recombination. These modified ES cells will permit efficient, selectable 'knock-in' of any form of rhodopsin, cDNA or genomic, wild-type or mutant, from any species, for a broad range of physiological studies. The investigator will "knock-in" modified human rhodopsin genes, designed to serve as sensitive cellular detectors of the effects of oligonucleotide treatments on rhodopsin gene expression, correction and knockout. For whole mouse studies, the investigator will use fusions of rhodopsin with green fluorescent protein (GFP). Oligonucleotide effects on transcription will be measured as a decrease in fluorescence intensity; oligonucleotide effects on gene correction and mutation will be measured by the appearance of GFP fluorescence starting with rhodopsin-GFP fusion constructs that are not expressed (GFP-). Cellular studies will precede whole animal experiments in order to test various treatment parameters. Although oligonucleotide-mediated effects on transcription can be measured readily, detection of recombination-based correction and mutational knockout requires specialized constructs, which the investigator will initially test at the selectable adenosine phosphoribosyl transferase (APRT) gene in CHO cells. A sensitive detection system will allow us to detect weak signals that can then be improved and optimized. Using CHO oligonucleotides (TFOs), RNA/DNA chimeric oligonucleotides and oligonucleotide analogues such as peptide nucleic acids (PNAs), for their ability to inhibit transcription, correct gene defects, and introduce mutations. These results will be used as the basis for developing treatment protocols in mice.